1. CD40
CD40 is an antigen having a molecular weight of 50 kDa which is present on the surface of cell membrane, and expressed in B cells, dendritic cells (DCs), some types of cancer cells, and thymic epithelial cells. CD40 is known to play an important role in proliferation and differentiation of B cells and DCs. CD40 was identified as an antigen expressed on the surface of human B cells (E. A. Clark et al., Proc. Natl. Acad. Sci. USA 83: 4494, 1986; and I. Stamenkovic et al., EMBO J. 8: 1403, 1989) and has been considered as a member of the TNF receptor family which includes low-affinity NGF receptors, TNF receptors, CD27, OX40 and CD30. Ligands (CD40Ls) to human and murine CD40s have been recently cloned and found to be membrane proteins type II and expressed in activated CD4+T cells. CD40L has been also found to introduce strong signals for activation into human or murine B cells.
In dendritic cells, CD40 has been observed to be more highly expressed than in B cells, and it has become clear that CD40 plays an important role in dendritic cells. Binding of CD40 to CD40L activates antigen presenting cells (APCs), that is, expresses costimulator molecules such as CD80 (B7-1) and CD86 (B7-2) or enhances production of IL-2 (Caux, C., et al.: Activation of human dendritic cells through CD40 cross-linking. J. Exp. Med., 180: 1263, 1994; and Shu, U., et al.: Activated T cells induce interleukin-12 production by monocyte via CD40 - CD40 ligand interaction. Eur. J. Immunol., 25: 1125, 1995). Dendritic cells have a strong antigen-presenting capacity and a strong capacity to activate helper T (Th) cells. Dendritic cells are also believed to control differentiation of naive Th cells into Th1 or Th2 cells. When peripheral blood monocytes, which are myeloid dendritic cells, are cultured in the presence of GM-CSF and IL-4, and matured by CD40L, the resulting matured dendritic cells (DC 1) can produce IL-12 in vitro, and stimulate and activate allogeneic naive Th cells to induce IFNγ-producing T cells (i.e., to promote their differentiation into Th1). This action is inhibited by anti-IL-12 antibody and hence may be effected via IL-12. On the other hand, when plasmacytoid T cells, which are present in lymphoid T regions and peripheral blood, are cultured in the presence of IL-3 and CD40 ligand, the resulting lymphoid dendritic cells (DC2) are shown to be unable to produce IL-12, and stimulate and activate allogeneic naive Th cells to induce IL-4-producing T cells, which indicates promotion of their differentiation into Th2. It is believed that Th1 cells are involved in activation of cellular immunity, while Th2 cells are associated with enhancement of humoral immunity as well as restriction of cellular immunity. When cytotoxic T cells (CTL) are activated with the help of Th1 cells, they may eliminate pathogens (a number of types of virus, listeria, tuberculosis bacteria, toxoplasma protozoa, etc.) growing in the cytoplasm and tumor cells.
Monoclonal anti-CD40 antibodies, which recognize CD40 expressed on the membrane surface, have been demonstrated to have different biological activities to B cells. Monoclonal anti-CD40 antibodies are generally classified into agonistic or antagonistic antibodies against the interaction between CD40 and CD40L.
2. Agonistic Antibodies
Agonistic antibodies are known to activate B cells. For instance, the anti-CD40 antibodies are reported to induce cell adhesion (Barrett et al., J. Immunol. 146: 1722, 1991; and Gordon et al., J. Immunol. 140: 1425, 1998), increase cell size (Gordon et al., J. Immunol. 140: 1425, 1998; and Valle et al., Eur. J. Immunol. 19: 1463, 1989), induce cell division of B cells activated only by an anti-IgM antibody, anti-CD20 antibody or phorbol ester (Clark and Ledbetter, Proc. NatI. Acad. Sci. USA 83: 4494, 1986; Gordon et al., LEUCOCYTE TYPING III. A. J. McMicheal ed. Oxford University Press. Oxford. p. 426; and Paulie et al., J. Immunol. 142: 590, 1989), induce cell division of B cells in the presence of IL4 (Valle et al., Eur. J. Immunol. 19: 1463, 1989; and Gordon et al., Eur. J. Immunol. 17: 1535, 1987), induce expression of IgE by cultured cells stimulated with IL-4 and deprived of T cells (Jabara et al., J. Exp. Med. 172: 1861, 1990; and Gascan et al., J. Immunol. 147: 8, 1991), induce expression of IgG and IgM by those cultured cells (Gascan et al., J. Immunol. 147: 8, 1991), secrete soluble CD23/FceRII from cells via IL-4 (Gordon and Guy, Immunol. Today 8: 39, 1987; and Cairns et al., Eur. J. Immunol. 18: 349, 1988), enhance expression of soluble CD23/FceRII on the cells via IL4 (Challa, A., Allergy, 54: 576, 1999), and promote IL-6 production (Clark and Shu, J. Immunol. 145: 1400, 1990). Furthermore, it is reported that addition of IL-4 and an anti-CD40 antibody to human primary culture B cells in the presence of CDw32+ adhesive cells led to establishment of cloned B cells derived therefrom (Bancherauet et al., Science 241: 70, 1991), and apoptosis of germinal center cells was inhibited through CD40 irrespective of whether its antigen receptor was active or inactive (Liu et al., Nature 342: 929, 1989). As described above, CD40 has been identified as antigen expressed on the surface of human B cells, and consequently, most of the isolated antibodies have been evaluated, as an index, mainly using their induction potency for proliferation and/or differentiation of human B cells, or their induction activity for cell death of cancer cells (Katira, A. et al., LEUKOCYTE TYPING V. S. F. Schlossossman, et. al. eds. p. 547. Oxford University Press. Oxford; W. C. Flansow et. al., LEUKOCYTE TYPING V. S. F. Schlossossman, et. al. eds. p. 555. Oxford University Press. Oxford; and J. D. Pound et. al., International Immunology, 11: 11, 1999).
The anti-CD40 antibody has been demonstrated to mature DC (Z. H. Zhou et. al., Hybridoma, 18: 471, 1999). Furthermore, the role of CD4 T cells in priming antigen-specific CD8 T cells was reported to be in activation of DC via CD40-CD40L signaling, and the anti-CD40 monoclonal antibody (mAb) has been found to be able to replace CD40 helper T cells in activation of dendritic cells (DC) (Shoenberger, S. P., et. al.: T-cell help for cytotoxic T lymphocytes is mediated by CD40-CD40L interactions. Nature, 480, 1998). Also, administration of an anti-CD40 antibody in mice has been found to be able to protect the animal body from CD40-expressing tumor cells as well as CD40-non-expressing tumor cells (French, R. R., et. al.: CD40 antibody evokes a cytotoxic T-cell response that eradicates lymphoma and bypasses T-cell help. Nature Medicine, 5, 1999).
Agonistic anti-CD40 antibodies are expected to be effective for treatment of infectious diseases, due to bacteria, virus, etc., cancer and others, based on their functions described above. Anti-CD40 antibodies with superior agonistic activities are described in WO 02/088186. The representative examples of those agonistic antibodies are KM341-1-19 and 2105 antibodies. The hybridoma KM341-1-19 producing the KM341-1-19 antibody and the hybridoma 2105 producing the 2105 antibody were submitted on 27, Sep. 2001 and 17, Apr. 2002, respectively, for international deposit under the Budapest Treaty, to International Patent Organisms Depositary, National Institute of Advanced Industrial Science and Technology (central 6, 1-1, Higashi 1, Tsukuba, Ibaraki, Japan). Their accession numbers are FERM BP-7759 (KM341-1-19) and FERM BP-8024 (2105).
3. Antagonistic Antibodies
Taking it in consideration, on the other hand, that CD40 plays an important role in immunologic responses, as aforementioned, it is expected that inhibition of binding of CD40 to its ligands would lead to development of therapeutic agents for immune suppression in organ transplantation and autoimmune diseases. Sawada, Hase and others have reported that the peripheral blood of patients suffering from Crohn's disease has a higher percentage of monocytes highly expressing CD40. However, such antibodies have not been well known yet as inhibit binding of CD40 to its ligands. Those inhibitory antibodies would be useful in functional analysis of CD40 and treatment of diseases requiring activation of CD40. Inhibitory antibodies to CD40 ligands are also suggested to be effective against diseases involving binding of CD40 to the CD40 ligands. However, CD40L was reported to be expressed in activated platelets (V. Henn et al., Nature 391: 591, 1998), and if an anti-CD40L antibody is used as a therapeutic agent, thrombus formation may occur reportedly (T. Kawai et al., Nat. Med. 6: 114, 2000). From this point of view, antibodies to CD40 are expected to be safer rather than anti-CD40L antibodies as therapeutic antibody agent to inhibit binding of CD40 to its ligands. Anti-CD40 antibodies would be required to inhibit binding of CD40L to CD40 and still not activate CD40 in themselves.
Such antagonistic anti-CD40 antibodies may be used for treatment of autoimmune diseases and suppression of immunologic rejections in transplantation of organs, bone marrow, etc., in view of their functions described above. Anti-CD40 antibodies with superior antagonistic activities are described in WO 02/088186. The representative example of those antagonistic antibodies is 4D11 antibody. The hybridoma 4D11 producing the 4D11 antibody was submitted on 27, Sep. 2001 for international deposit under the Budapest Treaty, to International Patent Organisms Depositary, National Institute of Advanced Industrial Science and Technology (central 6, 1-1, Higashi 1, Tsukuba, Ibaraki, Japan). The accession number is FERM BP-7758.
Patent Document 1 WO 02/088186